Pathogenesis of mucosal infections are often studied in monomicrobial settings that ignore the possible influence of other potentially pathogenic microbial flora present at the time of exposure. Urinary tract infection (UTI) is a common and sometimes persistent disease that can have life-threatening outcomes in susceptible populations. Women are at much higher risk of UTI than men, and are more likely to experience complications of infection during particular stages of the reproductive lifespan -- in pregnancy and after menopause. Abundant and diverse microbial populations exist in periurethral and nearby mucosal sites, therefore there is a high likelihood that bacterial exposures of the urinary tract are often polymicrobial. Uropathogenic E. coli (UPEC) is the most common cause of UTI, but often co-occurs with low levels of Gram positive bacteria in urine in a polymicrobial context. The presence of sub-diagnostic levels of Gram positives in urine is usually considered clinically insignificant; however, this assumption has never been tested experimentally. Group B Streptococcus (GBS) in a common commensal of the lower gastrointestinal and vaginal tracts and is commonly isolated from women with UPEC UTI or asymptomatic bacteriuria. This proposal describes the development of a novel murine model to study polymicrobial-host interactions in the urinary tract in mice. Proposed experiments use this model to investigste the impact of GBS on UPEC UTI outcomes. Preliminary data show that GBS has a striking impact on UPEC infection, altering the course, severity, and outcome of UTI. Ongoing cellular and molecular studies are defining specific host and bacterial factors involved in the polymicrobial dialogue. In this proposal, we aim to more fully understand the mechanisms driving alterations to the UPEC pathogenic cascade upon polymicrobial inoculation with GBS. In addition to paradigm-shifting implications for urinary pathogenesis, the proposed studies may uncover valuable predictors of risk in particular clinical settings.